This invention relates generally to spectroscopic methods. More particularly, in certain embodiments, the invention relates to an apparatus and methods for determining whether spectral data obtained from a region of a tissue sample are affected by one or more artifacts, such as glare, shadow, or an obstruction.
Spectral analysis may be used to diagnose disease in tissue. For example, spectral data may be obtained during a diagnostic procedure in which spectral scans are performed on the tissue of a patient. One such diagnostic procedure is an acetowhitening procedure, in which a chemical agent is applied to tissue and the response of the tissue is captured in a spectral scan at some point following the application of the agent. The chemical agent is used, for example, to enhance the detected difference between spectral data obtained from normal tissue and spectral data obtained from abnormal or diseased tissue.
Spectral measurements of tissue may be non-representative of the actual condition of the tissue when they are affected by one or more artifacts. Artifacts include lighting artifacts such as glare or shadow, and obstructions, such as blood, a speculum, a smoke tube, or other instruments used during the procedure. Artifacts may be located and determined using visual evidence of a region of tissue at the time of the procedure. However, there are currently no other suitable methods of determining whether spectral data obtained from a region of a tissue sample are affected by an artifact. Also, current methods of obtaining spectral data do not allow for the characterization of tissue in the event an artifact adversely affects the data.
The invention provides an apparatus and methods for obtaining redundant spectral data in order to compensate for artifacts that may be present in optical sample analysis. By illuminating a region of tissue with light incident to the region at more than one angle, it is possible to obtain redundant spectral data for the region. If one set of data for a region is adversely affected by an artifact such as glare, shadow, or an obstruction, then redundant data for the region, obtained using light incident to the region at a different angle, may be useful. The redundant data may be used to describe the region of tissue, unobscured by the artifact.
The invention comprises methods for determining if spectral data obtained from a sample region are affected by an artifact, and if so, whether or not redundant data may be used in place of the affected data. Embodiments of the invention comprise the use of metrics to determine whether an artifact is affecting the spectral data from a region of a tissue sample. Methods also comprise determining what kind of artifact is affecting the data from the region. These metrics involve computations using values of the spectral data corresponding to a the region of the tissue sample. Methods of the invention do not rely on any additional visual evidence of the tissue sample, such as human visual inspection, to determine the presence or absence of an artifact. In certain embodiments, the presence of an artifact is desired. However, in preferred embodiments, the presence of an artifact is not desired, for example, because the artifact adversely affects the spectral data.
If it is determined that an artifact has rendered unusable a given set of data for a region of the sample, then the redundant data corresponding to the region may be considered. Since the redundant data is obtained using light incident to the region at a different angle from that used to obtain the affected data, the artifact may not have affected the redundant data. Multiple sets of redundant data may be used in order to compensate for one or more artifacts. Preferred methods of the invention comprise determining whether redundant data are affected by an artifact or, alternatively, whether redundant data are unaffected by the artifact and representative of the unobscured tissue. If the set of redundant data is representative of an unobscured tissue, such data may be used in place of the affected data in characterizing the region or determining the condition of the region of tissue. As mentioned above, more than one redundant set of data may be obtained. Also, if more than one set of data is determined to be unaffected by an artifact, averages of the unaffected data may be used to characterize the region of tissue.
Although specific metrics were developed for application to the analysis of in vivo cervical tissue subject to artifacts such as glare, shadow, and obstructions, methods for developing analogous metrics are also disclosed as part of the invention. Such methods may be used to create metrics for the analysis of other types of tissue such as in vivo or ex vivo colorectral, gastroesophageal, urinary bladder, lung, skin tissue, and/or any tissue comprising epithelial cells, for example. These methods may be used to create metrics for tissues that are subject to other states of health and/or other types of artifacts, in addition to those discussed herein. The invention also comprises methods of determining computational metrics for use in applications employing different types of spectral data than those specifically discussed herein.
In most embodiments discussed herein, spectral data are obtained as a function of wavelength within a range of between about 360 nm and 720 nm. However, in some embodiments, the range of wavelengths is from about 190 nm to about 1100 nm. In the methods discussed herein, where a range of about 360 nm to about 720 nm is specified, a broader range within about 190 nm and about 1100 nm is alternately used for some embodiments.
In one aspect, the invention is directed to a method of determining a condition of a region of a tissue sample using two or more sets of spectral data, each set obtained using light incident to the region at a unique angle. The method comprises the steps of: obtaining a first set of spectral data corresponding to a region of a tissue sample using light incident to the region at a first angle; obtaining a second set of spectral data corresponding to the region using light incident to the region at a second angle; selecting at least one of the two sets that is representative of the region of the tissue sample; and determining a condition of the region of the tissue sample based at least in part on a portion of the representative data.
Both the first and the second sets of spectral data comprise reflectance spectral data in some embodiments. In other embodiments, at least one of the two sets of spectral data comprises fluorescence spectral data. In some embodiments, the method further comprises obtaining one or more additional sets of spectral data corresponding to a region of interest, each set using light incident to the region at a unique angle.
The condition to be determined may be a state of health. In one embodiment the state of health comprises at least one of the following conditions: normal squamous tissue, metaplasia, Cervical Intraepithelial Neoplasia Grade I (CIN I), Cervical Intraepithelial Neoplasia Grade II (CIN II), Cervical Intraepithelial Neoplasia Grade III (CIN III), carcinoma in-situ (CIS), and cancer. In some embodiments, the state of health is a combination of two or more of the conditions above, such as Cervical Intraepithelial Neoplasia Grade II or Grade III (CIN II/III).
In another aspect, the invention is directed to a method of determining whether spectral data obtained from a region of a tissue sample are affected by an artifact. The method comprises the steps of: obtaining a first set of spectral data corresponding to a region of a tissue sample using light incident to the region at a first angle; obtaining a second set of spectral data corresponding to the region using light incident to the region at a second angle; and determining whether the first set of data is affected by an artifact based at least in part on a portion of the data from each of the two sets.
Both the first and the second sets of spectral data comprise reflectance spectral data in some embodiments. In other embodiments, the method comprises obtaining a third set of spectral data comprising fluorescence spectral data.
The invention comprises methods of applying various computational metrics in determining whether or not spectral data are affected by an artifact. According to one embodiment, the method of determining whether spectral data are affected by an artifact comprises computing a difference between R1, a member of the first set of spectral data discussed above, and R2, a member of the second set of spectral data discussed above, and comparing the difference to a constant, where R1 and R2 correspond to at least approximately identical wavelengths. This difference is a percent difference in some preferred embodiments.
In some preferred embodiments, the method of determining whether spectral data are affected by an artifact comprises computing N differences, |R1(Xi)xe2x88x92R2(Xi)|, optionally weighting each of the N differences using at least one of R1(Xi) and R2(Xi), defining a maximum of a subset of the N optionally-weighted differences, and comparing the maximum to a first constant, where i=1 to N, N is an integer, Xi is a wavelength between about 360 nm and about 720 nm, R1(Xi) is a member of the first set of data corresponding to the wavelength Xi, and R2(Xi) is a member of the second set of data corresponding to the wavelength Xi.
The method of determining whether spectral data are affected by an artifact in some embodiments further comprises comparing R1(X1) to a second constant, where R1(X1) is a member of the first set of data corresponding to a wavelength X1 between about 409 nm and about 429 nm.
The method of determining whether spectral data are affected by an artifact in some embodiments further comprises comparing the quotient {(R1(X1)/R2(X1))/(R1(X2)/R2(X2))} to a second constant, where X1 is a wavelength between about 360 nm and about 720 nm, X2 is a wavelength between about 360 nm and about 720 nm, R1(X1) is a member of the first set of data corresponding to the wavelength X1, R2(X1) is a member of the second set of data corresponding to the wavelength X1, R1(X2) is a member of the first set of data corresponding to the wavelength X2, R2(X2) is a member of the second set of data corresponding to the wavelength X2. In one embodiment, X1 is a wavelength between about 566 nm and about 586 nm, and X2 is a wavelength between about 589 nm and about 609 nm. In one embodiments, the determining step further comprises comparing R1(X3) to a third constant, where R1(X3) is a member of the first set of data corresponding to a wavelength X3 between about 689 and about 709 nm. In another embodiment, X3 is between about 360 nm and about 720 nm. In yet another embodiment X3 is between about 409 nm and about 429 nm.
The method of determining whether spectral data are affected by an artifact in some embodiments further comprises comparing a value Q to a second constant, where Q is an approximate slope of a plot of {R1(Xi)/R2(Xi)} with respect to wavelength, over a subset of a wavelength range of about 360 nm to about 720 nm, Xi is a wavelength between about 360 nm and about 720 nm, R1(Xi) is a member of the first set of data corresponding to the wavelength Xi, and R2(Xi) is a member of the second set of data corresponding to the wavelength Xi.
In another embodiment, the method of determining whether spectral data are affected by an artifact further comprises comparing R1(X1) to a second constant and comparing R1(X1) to R2(X1), where R1(X1) is a member of the first set of data corresponding to a wavelength X1 between about 360 nm and about 720 nm, and R2(X1) is a member of the second set of data corresponding to the wavelength X1.
The method of determining whether spectral data are affected by an artifact in another embodiment further comprises comparing R1(X1) to a second constant and comparing R1(X1) to R2(X1), where R1(X1) is a member of the first set of data corresponding to a wavelength X1 between about 489 nm and about 509 nm, and R2(X1) is a member of the second set of data corresponding to the wavelength X1.
According to some embodiments, the method of determining whether spectral data are affected by an artifact comprises comparing R1(X1) to a constant, where R1(X1) is a member of the first set of data corresponding to a wavelength X1 between about 409 nm and about 429 nm. In one embodiment, the determining step further comprises comparing a value Q to a second constant, where the value Q is an approximate slope of a plot of {R1(Xi)/R2(Xi)} with respect to wavelength, over a subset of a wavelength range of about 576 nm to about 599 nm, Xi is a wavelength between about 360 nm and about 720 nm, R1(Xi) is a member of the first set of data corresponding to the wavelength Xi, and R2(Xi) is a member of the second set of data corresponding to the wavelength Xi.
In some embodiments, the method of determining whether spectral data are affected by an artifact comprises comparing the quotient R1(Xi)/R1(X2) to a constant, where R1(X1) is a member of the first set of data corresponding to a wavelength X1 between about 360 nm and about 720 nm, and R1(X2) is a member of the first set of data corresponding to a wavelength X2 between about 360 nm and about 720 nm. In one embodiment, X1 is a wavelength between about 489 nm and 509 nm and X2 is a wavelength between about 533 nm and about 553 nm.
According to one embodiment, the method of determining whether spectral data are affected by an artifact comprises comparing R1 to a first constant and comparing R2 to a second constant, where R1 is a member of the first set of data corresponding to a wavelength between about 489 nm and about 509 nm and R2 is a member of the second set of data corresponding to a wavelength between about 489 nm and about 509 nm.
The artifact comprises a lighting artifact in some embodiments. The lighting artifact comprises glare and/or shadow in some embodiments. In other embodiments, the artifact comprises an obstruction. An obstruction comprises blood, mucus, a speculum, and/or a smoke tube in these embodiment. There may be both lighting artifacts and obstruction artifacts in a given embodiment.
According to one embodiment, the tissue sample comprises cervical tissue. In some embodiments, the tissue sample contains epithelial cells as tissue components. The tissue sample comprises at least one of a group consisting of cervical, colorectal, gastroesophageal, urinary bladder, lung, and skin tissue in some embodiments.
In another aspect, the invention is directed to a method of determining whether spectral data corresponding to a region of a tissue sample is affected by an artifact using two sets of reflectance spectral data and one set of fluorescence spectral data. The method comprises the steps of: obtaining a first set of reflectance spectral data corresponding to a region of a tissue sample using light incident to the region at a first angle; obtaining a second set of reflectance spectral data corresponding to the region using light incident to the region at a second angle; obtaining a set of fluorescence spectral data corresponding to the region; and determining whether any of the data is affected by an artifact based at least in part on at least one of the following: a subset of the first set of reflectance spectral data, a subset of the second set of reflectance spectral data, and a subset of the set of fluorescence spectral data. In one embodiment, the determining step comprises comparing F to a constant, where F is a member of the set of fluorescence spectral data corresponding to a wavelength between about 469 nm and about 489 nm.
In another aspect, the invention is directed to methods of determining a spectral characteristic of an artifact. These include methods of determining computational metrics used to judge whether spectral data obtained from a region are affected by an artifact. A preferred method comprises the steps of: (a) at each of a first plurality of regions of tissue, obtaining a first set of reflectance spectral data known to be affected by a given artifact; (b) at each of a second plurality of regions of tissue, obtaining a second set of reflectance spectral data known not to be affected by the artifact; and (c) determining a spectral characteristic of the artifact based at least in part on the first and second sets of reflectance spectral data.
The method of determining a spectral characteristic in some embodiments comprises locating a wavelength at which there is a maximum difference between a mean of one or more members of the first set of reflectance spectral data corresponding to the wavelength and a mean of one or more members of the second set of reflectance spectral data corresponding to the wavelength, relative to a variation measure.
In some embodiments, the method of determining a spectral characteristic comprises computing N differences, |xcexci(Aj(Xi))xe2x88x92xcexci(Bk(Xi))|, and defining a maximum of a subset of the N differences, where i=1 to N, N is an integer, Xi is a wavelength between about 360 nm and about 720 nm, j=1 to M1, M1 is an integer, Aj(Xi) represents one of M1 members of the first set of reflectance spectral data corresponding to the wavelength Xi, k=1 to M2, M2 is an integer, Bk(Xi) represents one of M2 members of the second set of reflectance spectral data corresponding to the wavelength Xi, xcexci(Aj(Xi)) is a mean of the M1 members of the first set of data corresponding to the wavelength Xi, and xcexc(Bk(Xi)) is a mean of the M2 members of the second set of data corresponding to the wavelength Xi.
According to some embodiments, the method of determining a spectral characteristic comprises computing N quotients, [|xcexci(Aj(Xi))xe2x88x92xcexci(Bk(Xi))|/{"sgr"i2(Aj(Xi))+"sgr"i2(Bk(Xi))}0.5], and defining a maximum of a subset of the N quotients, where i=1 to N, N is an integer, X, is a wavelength between about 360 nm and about 720 nm, j=1 to M1, M1 is an integer, Aj(Xi) represents one of M1 members of the first set of reflectance spectral data corresponding to the wavelength Xi, k=1 to M2, M2 is an integer, Bk(X1) represents one of M2 members of the second set of reflectance spectral data corresponding to the wavelength Xi, xcexci(Aj(Xi)) is a mean of said M1 members of the first set of data corresponding to the wavelength Xi, xcexci(Bk(Xi)) is a mean of the M2 members of the second set of data corresponding to the wavelength Xi"sgr"i(Aj(Xi)) represents a standard deviation of the M1 members of the first set of data corresponding to the wavelength Xi, and "sgr"i(Bk(Xi)) represents a standard deviation of the M2 members of the second set of data corresponding to the wavelength Xi.
The method of determining a spectral characteristic in some embodiments comprises computing N quotients, [|xcexci(Aj(X1i)/Aj(X2i))xe2x88x92xcexci(Bk(X1i)/Bk(X2i))|/{"sgr"i(Aj(X1i)/Aj(X2i))+"sgr"i2(Bk(X1i)/Bk(X2i))}0.5], and defining a maximum of a subset of the N quotients, where i=1 to N, N is an integer, X1, is a wavelength between about 360 nm and about 720 nm, X2, is a wavelength between about 360 nm and about 720 nm, j=1 to M1, M1 is an integer, Aj(X1,) represents one of M1 members of the first set of reflectance spectral data corresponding to the wavelength X1,, Aj(X2i) represents one of M1 members of the first set of reflectance spectral data corresponding to the wavelength X2i, k=1 to M2, M2 is an integer, Bk(X1i) represents one of M2 members of the second set of reflectance spectral data corresponding to the wavelength X1i, Bk(X2i) represents one of M2 members of the second set of reflectance spectral data corresponding to the wavelength X2ixcexci(Aj(X11)/Aj(X2i)) is a mean of M1 quotients Aj(X1i)/Aj(X2i) for j=1 to M1, xcexci(Bk(X1i)/Bk(X2i)) is a mean of M2 quotients Bk(X1i)/Bk(X2i) for k=1 to M2, "sgr"i(Aj(X1i)/Aj(X2i)) represents a standard deviation of the M1 quotients Aj(X1i)/Aj(X2i), and "sgr"i(Bk(X1i)/Bk(X2i)) represents a standard deviation of the M2 quotients Bk(X1i)/Bk(X2i).
In another aspect, the invention is directed to a method of determining a characteristic of a region of a tissue sample by obtaining at least two sets of reflectance spectral data, each using light incident to the region at a different angle, and eliminating data that is adversely affected by an artifact. The method comprises the steps of: (a) obtaining a first set of reflectance spectral data corresponding to a region of a tissue sample using light incident to the region at a first angle; (b) obtaining a second set of reflectance spectral data corresponding to the region using light incident to the region at a second angle; (c) determining whether at least one of the first set of reflectance data and the second set of reflectance data is affected by an artifact based at least in part on a subset of the first set of reflectance data and a subset of the second set of reflectance data; (d) rejecting at least one member of at least one of the first set of reflectance data and the second set of reflectance data determined in step (c) to be affected by the artifact; (e) determining a characteristic of the region of the tissue sample based at least in part on at least one member of at least one of the first set of reflectance data and the second set of reflectance data not rejected in step (d).
In some embodiments, the method further comprises obtaining a set of fluorescence spectral data corresponding to the region, and step (e) comprises determining the condition of the region of the tissue sample based at least in part on at least one member of at least one of the first set and the second set of reflectance data and at least one member of the set of fluorescence spectral data.
Although certain embodiments of the invention are specifically described with respect to fluorescence spectral data and/or reflectance (backscatter) spectral data, these methods may be adapted for use with other kinds of optical signal data that may be affected by artifacts, including Raman, infrared, video signal data, and combinations thereof.